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Irradiation is an established method of sterilization for pharmaceutical products. Radiation sterilization can be achieved with gamma rays, electron beams, and X-rays. Each of these techniques has its advantages and disadvantages. The author describes these methods, the ways to find the correct sterilization doses, and the regulatory and safety concerns about irradation sterilization.
Drug makers have sterilized pharmaceuticals by gamma irradiation for more than 40 years. High-energy gamma irradiation is used mainly in the healthcare industries to sterilize disposable medical devices. Over the years, however, the number of radiation-sterilized pharmaceuticals has gradually increased. Pharmaceutical companies now radiation sterilize drugs such as ophthalmic preparations, topical ointments, veterinary products, and parenterals. Regulatory pressure to adopt terminal-sterilization processes has promoted radiation sterilization.
Radiation sterilization may be performed using either gamma rays from a radioisotope source (usually cobalt-60) or electron-beam or X-ray irradiation. Gamma-ray irradiation, however, is by far the more common method.
Like all methods of sterilization, irradiation involves a compromise between inactivation of the contaminating microorganisms and damage to the product being sterilized. The imparted energy, in the form of gamma photons or electrons, does not always differentiate between molecules of the contaminating microorganism and those of the pharmaceutical substrate.
The interaction between high-energy gamma radiation and matter forms ion pairs by ejecting electrons, leading to free-radical formation and excitation. The free radicals are extremely reactive because each has an unpaired electron on one of its outer orbitals. Free-radical reactions may involve gas liberation, double-bond formation and scission, exchange reactions, electron migration, and cross-linking. In fact, any chemical bond may be broken and any potential chemical reaction may take place. In crystalline materials, this may result in vacancies, interstitial atoms, collisions, thermal spurs, and ionizing effects. Polymerization is a particularly common result in unsaturated compounds. In microorganisms, radiation-induced damage may express itself in various biological changes that may lead to cell death. Although DNA generally is considered the major subject of cellular damage, membrane damage also may contribute significantly to reproductive-cell death. In solutions, a molecule may receive energy directly from the incident radiation (the "direct effect") or, in aqueous solutions such as parenterals, by the transfer of energy from the radiolysis products of water (e.g., hydrogen and hydroxyl radicals and the hydrated electron) to the solute molecule (the "indirect effect"). In dilute solutions, most of the energy is imparted to the water, as is the case with many parenteral solutions. The indirect radiation effect therefore would account for most of the resulting possible radiation damage.
The process of radiation-induced damage by electrons is similar to that for gamma photons. In electron irradiation, the high-energy electrons produced outside the target molecule ionize the molecular species as they pass through the medium and release their energy. The ionization process leads to the production of secondary electrons with a range of energies capable of breaking bonds in the medium near the ionization event. The high-energy electrons usually are produced either by accelerating them across a large drop in potential in a direct-current machine or by a linear or circular electron accelerator.
X-rays are electromagnetic photons emitted when high-energy electrons strike any material. X-rays therefore can be produced by an electron accelerator. X-ray sterilization is not as fast as electron-beam irradiation. Since electron-beam and X-ray machines are powered electrically, the handling, shipping, and disposal of radioisotopes is not necessary. A disadvantage of electron-beam irradiation is its low penetration power, although more modern machines have overcome this problem. X-ray machines may penetrate even more than gamma-ray machines.
The chapters about gamma-radiation and electron-beam sterilization in the Encyclopedia of Pharmaceutical Technology contain general reviews of radiation sterilization (1, 2).
Contract sterilizers usually perform irradiation (1, 3). Though the contract sterilizer usually assumes many process-validation duties, the drug manufacturer bears final responsibility for the product's sterility. The contract sterilizer essentially is responsible for guaranteeing the delivered radiation dose.
The effect of radiation on pharmaceuticals
Any processing such as sterilization in the manufacture of a pharmaceutical product must cause minimal degradation. This requirement applies to radiation processing. Data on the feasibility of irradiating final pharmaceutical products (parenteral products in particular), active ingredients, or excipients can be obtained from the scientific literature. Reviews on the effects of gamma and electron-beam irradiation are readily available (4–17). Although many of the cited investigations offer only a superficial examination of the irradiated drugs, the reported data give useful insights into the overall radiation stability of these products and indicate whether more extensive testing of the products should be undertaken.
It is necessary to examine each new compound to assess its radiation stability, even though data may be available for closely related compounds. A thorough knowledge of radiation chemistry would be necessary to infer the behavior of one compound from another. Furthermore, with a formulated medication, the stability of an individual component may change when irradiated as part of the product.
Although sterilization doses of radiation usually are on the order of 25 kGy, a higher dose such as 50 kGy is useful for feasibility studies to indicate the type of radiolytic decomposition that may be expected at sterilization-dose levels.
Several analytical tools should be used to detect radiation-induced degradation. Each technique usually reveals a change in a specific moiety of the irradiated molecule, and it is therefore essential to examine all generated data to discover the extent of degradation. Stability-indicating assays should be used wherever possible. As with all stability studies, assays should be carried out during an extended time period to reveal the product's long-term stability. Accelerated aging may be undertaken under conditions recommended by the appropriate regulatory authority such as the US Food and Drug Administration.
Even when radiolysis products are within acceptable compendial limits, one must establish conclusively that the products formed cause no adverse effects at the concentrations found. Radiolysis products, however, generally are not unique to irradiation. It often suffices to show that radiolysis products are the same as those found when the drug is subjected to other sterilization procedures and occur at similar concentrations.
Irradiation of water. When discussing the irradiation of pharmaceuticals, particularly parenterals, the effects of irradiation on water must be considered. The formation of the various radiolysis products of water reflects the complexity of this "simple" pharmaceutical system. While none of the radical species formed are stable, they may react with the product's active ingredient, excipients, or both. The only resulting final products, however, are H2O, H2O2, and H2. Studies of the feasibility of radiosterilizing water in various container materials have been carried out in several laboratories (18–20).
A study of the effect of irradiated water on oxidation-susceptible drugs such as beta-lactam antibiotics showed that the drugs generally are not degraded. Furthermore, levels of H2O2 produced are below toxic levels (21).
Irradiation of powders for injection. The author's investigations of the radiation sterilization of parenterals have focused on powders of the beta-lactam group of antibiotics (essentially penicillins and cephalosporins). The rationale for these studies was beta-lactams' susceptibility to hydrolysis, particularly at elevated temperatures, which precludes the sterilization of parenteral solutions by conventional methods such as autoclaving. The necessity of sterilizing powders for injection by costly and highly demanding aseptic processes makes sterilization by irradiation most desirable.
Other applications. Other specific applications of irradiation to sterilize parenterals include complex drug-delivery systems and multicomponent parenteral units such as those in which the drug powder and solvent are compartmentalized until administration. Irradiation also may be used to decontaminate problematic raw materials such as active ingredients or pharmaceutical adjuncts used to manufacture parenterals.
Minimizing radiolysis. The formation of radiolysis products sometimes can be reduced. For example, irradiation may be undertaken in anoxia, at low temperatures, or by incorporating suitable additives if the degradation pathways are known. Of course, additives must not be toxic or interfere with the efficacy of the drug. They may include energy-transfer systems, -SH containing molecules, scavengers of radiolysis products of water, or reagents that convert radiolysis products to the parent compound. One example of such a radiation-tailored formulation is urea broth, which is used to identify Proteus species and differentiate it from other Gram-negative intestinal bacteria (22).
Radiolysis sometimes may be reduced by using electron-beam irradiation rather than gamma irradiation. The dose rate may be an important factor in electron-beam irratiation. Although no general rule exists, many drugs show less breakdown at a higher dose rate, i.e., with electron-beam irradiation. Reduced breakdown may result from the consumption of all the oxygen (which generally increases radiation damage) and the completion of sterilization before oxygen can be replenished. The process also may last too short a time to produce long-lived free radicals that could increase radiation-induced damage. On the other hand, the high dose rate could cause increased damage in some cases because of the high concentration of gamma photons close to the substrate.
Packaging materials. The radiation stability of packaging and container materials must never be overlooked when considering radiation compatibility. Lists of radiation-compatible packaging materials are readily available (1, 6, 22–25).
Validation of radiation sterilization
Validation of the radiation-sterilization process is an integral aspect of good manufacturing practice. It comprises installation qualification (IQ), operational qualification (OQ), performance qualification (PQ), materials compatibility, selection of sterilization dose, and routine process control. These components of validation relate either to the irradiation facility itself or the product being irradiated.
IQ, or irradiator commissioning, ensures that the irradiator has been supplied and installed in accordance with its specifications.
The essential parameter that must be controlled in radiation sterilization is measurement of radiation dose. Measurement is achieved using dosimeters, which are chemical or physical systems that respond quantitatively to absorbed radiation doses.
OQ demonstrates that the installed irradiator can operate and deliver appropriate radiation doses within defined acceptance criteria. PQ is essentially dose mapping. During dose mapping, the location and magnitude of the minimum and maximum delivered doses must be identified.
Radiation units. The absorbed radiation dose generally is expressed in rads (radiation absorbed doses). One rad is equivalent to an absorbed energy of 100 erg/g of material. The currently used SI unit for radiation-absorbed doses, however, is the gray, which is equivalent to an energy absorption of 1 joule/kg. One gray is equivalent to 100 rad, and 25 kGy, a common radiation dose for sterilization, is equivalent to 2.5 Mrad.
Determination of sterilization dose
An integral part of sterilization-process validation is the determination of a radiation dose for sterilization. Any deviation from the selected dose could either compromise the sterility of the product or damage the product.
A radiation dose of 25 kGy (2.5 Mrad) generally is accepted as suitable for sterilization purposes. This dose was chosen according to the radiation resistance of the bacterial spores of Bacillus pumilus. Today, the choice of radiation dose is based on the presterilization microbial contamination, or bioburden, and the desired sterility assurance level (SAL) of the product. Such considerations are based in part on extensive studies of the effects of substerilization doses on different microbial populations (27, 28). SAL is defined as the probability of a single viable microorganism occurring on a product following sterilization. SAL normally is expressed as 10–n. While the majority of authorities give n a value of 6, FDA does allow values of less than 6 for noninvasive products.
Most regulatory authorities expect radiation sterilization doses to be selected according to one of the methods of the International Organization for Standardization (ISO) standard, ISO 11137-2:2006 (Sterilization of Health Care Products—Radiation—Part 2: Establishing the Sterilization Dose). ISO 11137:2006 is published in three sections that discuss radiation sterilization, establishing the sterilization dose for radiation sterilization, and dosimetric aspects of radiation sterilization. The dose-setting methods described in the AAMI–ISO standards owe much to the ideas first presented by Tallentire and his colleagues (26).
The first ISO method, designated Method 1, is certainly the most common method used for dose selection for sterilization. The method requires the average microbial contamination of representative samples of the product to be determined. Note that the microbial population's radiation resistance is not determined. Dose setting is based on manufacturers' data about the resistance of microbial populations. The distribution of the chosen resistance is assumed to represent a more severe challenge than that presented by the natural bioburden of the article to be sterilized. The assumption is verified experimentally by irradiating 100 samples at a given verification dose and is accepted if no more than two contaminated samples remain. The sterilizing dose, which is appropriate for the average bioburden per sample and the desired SAL for the product, is then read from a table.
Method 2 does not entail the measurement of the bioburden. It relies on a protocol for a series of incremental-dose experiments to establish a dose at which approximately one in a hundred samples will be nonsterile. A sterilization dose then is established by extrapolation from this 10–2 sterility level using a dose-resistance factor calculated from observations of the incremental-dose experiments that characterize the remaining microbial resistance. This resistance is estimated from the lowest incremental dose at which at least one sample is sterile and from the dose at which the surviving population is estimated to be 0.01 microorganisms per sample.
The Verification-Dose (VDmax) Method, a relatively new method, is included in the current AAMI–ISO guidelines specifically to substantiate a 25-kGy dose. This method was officially introduced as an AAMI Technical Information Report (28) and is now part of ISO 11137-2: 2006. Kowalski and Tallentire proposed this method to substantiate a 25-kGy dose (29). The method is similar to dose-setting Method 1 because it requires a determination of bioburden and a verification dose experiment.
By substantiating a 25-kGy dose, this method verifies that the bioburden on the product is less radiation-resistant than a microbial population of maximal resistance, consistent with an SAL of 10–6 at 25 kGy. Verification is undertaken at an SAL of 10–1. Ten items are irradiated during the verification-dose experiment. The dose corresponding to this SAL (VDmax) reflects both the magnitude of the bioburden and the associated maximal resistance. If no more than one of ten sterility tests is positive, a 25-kGy sterilization dose is substantiated.
ISO also allows 25-kGy doses to be substantiated using Methods 1 and 2. The new ISO guidelines (ISO 11137-2:2006) allow dose-setting by other methods that provide assurance equivalent to that of the above ISO methods in achieving the specified sterility requirements. All ISO methods require periodic audits to confirm the appropriateness of the sterilization dose.
Other regulatory considerations
Although radiation sterilization has appeared in the United States Pharmacopeia since 1965, the US Food and Drug Administration regards a radiation-sterilized drug as a new product. Manufacturers must submit new drug applications and prove the products' safety.
In the United Kingdom, sterilization by ionizing radiation has been a recognized method since 1980, when the Ministry of Health agreed to accept materials exposed to a radiation dose of 25 kGy. Medicines controlled under the UK Medicines Act of 1968 are subjected to individual assessment by the Medicines and Healthcare Products Regulatory Agency's Committee on Safety of Medicines. This committee requires proof of sterility, proof that the drug's potency is unaffected by the process, and proof that degradation products are not harmful.
Although the British Pharmacopoeia recognizes gamma irradiation as a suitable sterilization process, the manufacturer must prove that no product degradation has taken place.
Most European countries allow pharmaceuticals to be radiation sterilized, provided that authorization has been obtained from the appropriate health authorities.
While the author does not believe that irradiation normally should replace traditional methods of sterilization for common, large-volume parenterals, irradiation should be considered seriously to sterilize powders for injection and small-volume parenterals that currently are sterilized by nonterminal sterilization processes.
Geoffrey P. Jacobs, PhD, is managing director of Dr. Geoffrey P. Jacobs Associates, PO Box 16352, Jerusalem 91162, Israel, tel. +972 2 6422227, fax +972 2 6432372, firstname.lastname@example.org
Keywords: irradiation, product stability, sterilization, validation.
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